Collective Responses to Malaria Vaccination

总结 总体项目目标是确定减毒疫苗接种免疫应答的关键特征， 恶性疟原虫子孢子（aPfSPZ），其导致绝育免疫。我们假设这 免疫是由于细胞和体液反应，特异性Pf抗原（Ag），提出了在 感染的肝脏阶段（LS）。我们提出了一个详细的纵向分析先天性和适应性免疫 使用尖端的免疫学分析，相关信息学和系统免疫学的反应。我们 将确定aSPZ疫苗诱导的与疟疾保护相关的细胞和体液反应 初治和有疟疾经历的受试者。互补的正交分析工具和方法将 鉴定相关免疫细胞亚群、Ag受体（AR）库、谱系和保护性免疫细胞的特异性。 艾格斯免疫细胞和产物（包括抗体）的功能特征与 保护将得到确认和验证。目标1：鉴定aSPZ疫苗 诱导与保护相关的细胞和体液反应。无偏倚和Ag特异性细胞 分析将表征与疫苗接种和CHMI相关的保护反应的动力学。后续 分析将通过质谱细胞术（CyTOF）描绘鉴定的Ag特异性适应性和先天性细胞应答， 多重pMHC/Ag多聚体染色和多组学单细胞测序以确定保护- 相关的基因和蛋白质表达特征和功能性抗体反应，这将被扩展 后来的目标。目的2：确定保护相关的T和B细胞Ag受体（AR）库在关键时间后， 疫苗和CHMI。将对来自T和B细胞的天然配对和单个AR链进行测序， 确定库属性并鉴定疫苗诱导的Pf Ag特异性AR。细胞转录组谱 和Ag特异性将通过联合的单细胞和TCR/BCR序列分析来联系。新型AR-Ag和 通过酵母展示方法鉴定抗体特异性。这一目标将鉴定T和B细胞AR， 与保护相关的抗体Ag特异性。目标3：确定 提供保护的细胞和抗体反应。分选的反应性T细胞将通过Ag特异性免疫组织化学法进行测定。 基于CyTOF的细胞内细胞因子染色（ICS）、激活诱导标记物（AIM）和单细胞测序 基于多组学分析。单克隆抗体的生物物理和功能分析将用于 评估B细胞在保护中的作用。综合项目数据将经过综合多因素 生物信息学分析，管理和共享使用已建立的平台。总的来说，该项目将确定关键 对aSPZ Pf疫苗接种的复杂抗原组成的复杂免疫应答的特征 以及与保护相关的效应过程，并提供一组保护性免疫相关物， 以指导疫苗开发。

SUMMARY The overall project goal is to identify key characteristics of immune responses to vaccination with attenuated Plasmodium falciparum sporozoites (aPfSPZs) which results in sterilizing immunity. We hypothesize that this immunity is due to cellular and humoral responses to specific to Pf antigens (Ags) that are presented during the liver stage (LS) of infection. We propose a detailed longitudinal analysis of innate and adaptive immune responses using cutting-edge immunological analyses, associated informatics, and systems immunology. We will identify aSPZ vaccine-induced cellular and humoral responses that correlate with protection in malaria naïve and malaria experienced subjects. Complementary orthogonal analytical tools and approaches will identify relevant immune cell subsets, Ag receptor (AR) repertoires, lineages, and specificities to protective Ags. The functional characteristics of the immune cells and products, including antibodies, that correlate with protection will be identified and validated. This will be accomplished by: Aim 1: Identifying aSPZ vaccine induced cellular and humoral responses that are associated with protection. Unbiased and Ag specific cellular profiling will characterize the kinetics of protection associated responses to vaccination and CHMI. Follow-on analyses will profile identified Ag-specific adaptive and innate cellular responses by mass cytometry (CyTOF), multiplexed pMHC/Ag multimer staining, and multi-omic single cell sequencing to determine protection- associated gene and protein expression signatures and functional antibody responses, which will be extended in later aims. Aim 2: Identify protection associated T and B cell Ag receptor (AR) repertoires at key times after vaccinations and CHMI. Natively paired and single ARs chains from T and B cells will be sequenced to determine repertoire attributes and identify vaccine induced Pf Ag specific ARs. Cellular transcriptomic profiles and Ag specificities will be linked by joint single cell and TCR/BCR sequence analyses. Novel AR-Ag and antibody specificities will be identified by yeast display methods. This aim will identify T and B cell ARs and antibody Ag specificities that are correlated with protection. Aim 3: Identifying functional characteristics of cellular and antibody responses that confer protection. Sorted reactive T cells will be assayed by Ag-specific CyTOF-based intracellular cytokine staining (ICS), activation-induce marker (AIM), and single-cell sequence based multi-omics analyses. The biophysical and functional analyses of monoclonal antibodies will be used to assess B cell roles in protection. The comprehensive project data will undergo integrated multi-factorial bioinformatic analysis, curation and sharing using established platforms. Overall, this project will identify key characteristics of the intricate immune responses to the complex antigenic composition of aSPZ Pf vaccination and the effector processes that correlate with protection and provide a set of immune correlates of protective immunity against malaria to guide vaccine development.